TLR ligands are promising candidates for the development of novel vaccine adjuvants that can elicit protective immunity against emerging infectious diseases. Adjuvants have been used most frequently to increase the quantity of an immune response. However, the quality of a T cell response can be more important than its quantity. Stimulating certain pairs of TLRs induces a synergistic response in terms of activating dendritic cells and eliciting/enhancing T cell responses through clonal expansion, which increases the number of responding T cells. Here, we have found that utilizing ligands for 3 TLRs (TLR2/6, TLR3, and TLR9) greatly increased the protective efficacy of vaccination with an HIV envelope peptide in mice when compared with using ligands for only any 2 of these TLRs; surprisingly, increased protection was induced without a marked increase in the number of peptide-specific T cells. Rather, the combination of these 3 TLR ligands augmented the quality of the T cell responses primarily by amplifying their functional avidity for the antigen, which was necessary for clearance of virus. The triple combination increased production of DC IL-15 along with its receptor, IL-15Rα, which contributed to high avidity, and decreased expression of programmed death–ligand 1 and induction of Tregs. Therefore, selective TLR ligand combinations can increase protective efficacy by increasing the quality rather than the quantity of T cell responses.

The mucosa of the large intestine mucosa is an effective vaccination site for induction of protective mucosal immunity against rectal or vaginal viral infection but clinically is impractical. Here Qing Zhu et al. have developed an oral delivery system that encapsulates vaccine into pH-dependent poly(DL-lactic-co-glycolic acid) (PLGA) nanoparticles coated with Eudragit to protect against the low pH and enzymatic destruction of the stomach. This approach was shown to selectively target the large intestine in mice and induce antigen-specific T and B cell responses similar to that observed with intracolorectal vaccination. Both rectal and vaginal mucosal surfaces serve as transmission routes for pathogenic microorganisms. Vaccination through large intestinal mucosa, previously proven protective for both of these mucosal sites in animal studies, can be achieved successfully by direct intracolorectal (i.c.r.) administration, but this route is clinically impractical. Oral vaccine delivery seems preferable but runs the risk of the vaccine's destruction in the upper gastrointestinal tract. Therefore, we designed a large intestine–targeted oral delivery with pH-dependent microparticles containing vaccine nanoparticles, which induced colorectal immunity in mice comparably to colorectal vaccination and protected against rectal and vaginal viral challenge. Conversely, vaccine targeted to the small intestine induced only small intestinal immunity and provided no rectal or vaginal protection, demonstrating functional compartmentalization within the gut mucosal immune system. Therefore, using this oral vaccine delivery system to target the large intestine, but not the small intestine, may represent a feasible new strategy for immune protection of rectal and vaginal mucosa.

Adjuvant effects on innate as well as adaptive immunity may be critical for inducing protection against mucosal HIV and simian immunodeficiency virus (SIV) exposure. We therefore studied effects of Toll-like receptor agonists and IL-15 as mucosal adjuvants on both innate and adaptive immunity in a peptide/poxvirus HIV/SIV mucosal vaccine in macaques, and made three critical observations regarding both innate and adaptive correlates of protection: ( i ) adjuvant-alone without vaccine antigen impacted the intrarectal SIVmac251 challenge outcome, correlating with surprisingly long-lived APOBEC3G (A3G)-mediated innate immunity; in addition, even among animals receiving vaccine with adjuvants, viral load correlated inversely with A3G levels; ( ii ) a surprising threshold-like effect existed for vaccine-induced adaptive immunity control of viral load, and only antigen-specific polyfunctional CD8 ⁺ T cells correlated with protection, not tetramer ⁺ T cells, demonstrating the importance of T-cell quality; ( iii ) synergy was observed between Toll-like receptor agonists and IL-15 for driving adaptive responses through the up-regulation of IL-15Rα, which can present IL-15 in trans , as well as for driving the innate A3G response. Thus, strategic use of molecular adjuvants can provide better mucosal protection through induction of both innate and adaptive immunity.

Effective SARS-CoV-2 vaccines are urgently needed. Although most vaccine strategies have focused on systemic immunization, here we compared the protective efficacy of 2 adjuvanted subunit vaccines with spike protein S1: an intramuscularly primed/boosted vaccine and an intramuscularly primed/intranasally boosted mucosal vaccine in rhesus macaques. The intramuscular-alum–only vaccine induced robust binding and neutralizing antibody and persistent cellular immunity systemically and mucosally, whereas intranasal boosting with nanoparticles, including IL-15 and TLR agonists, elicited weaker T cell and Ab responses but higher dimeric IgA and IFN-α. Nevertheless, following SARS-CoV-2 challenge, neither group showed detectable subgenomic RNA in upper or lower respiratory tracts versus naive controls, indicating full protection against viral replication. Although mucosal and systemic protective mechanisms may differ, results demonstrate both vaccines can protect against respiratory SARS-CoV-2 exposure. In summary, we have demonstrated that the mucosal vaccine was safe after multiple doses and cleared the input virus more efficiently in the nasal cavity and thus may act as a potent complementary reinforcing boost for conventional systemic vaccines to provide overall better protection.